Aggregatibacter (Actinobacillus) actinomycetemcomitans is a Gram-negative coccobacillus of the Pasteurellaceae family. It is implicated in periodontitis, a common low-grade bacterial infection, but it can also cause non-oral infections. The main aim of this project was to identify and characterize in A. actinomycetemcomitans novel cell surface components bearing virulence potential that could contribute to systemic immunoinflammatory burden. We first established and evaluated a method for preparing homogeneous cell suspensions of autoaggregating clinical isolates of A. actinomycetemcomitans. The chosen method is based on a gradual dispersion of bacterial colonies into solution, which generated homogeneous suspensions without losing cell viability or fimbriation. When sera from two patients with A. actinomycetemcomitans-associated infections were used to probe A. actinomycetemcomitans outer membrane protein (OMP) preparations in western blot, strong reactions were found at 17 kDa. Interestingly, antiserum against CsgA, a major subunit of Eschirichia coli curli, also reacted with A. actinomycetemcomitans OMP preparations at 17 kDa size, that is the size of E. coli CsgA, suggesting antigenic crossreactivity. The 17 kDa A. actinomycetemcomitans OMP was subsequently identified as peptidoglycan-associated lipoprotein (PAL; AaPAL) by using immunoproteomics methods. Studies on the pal gene and its gene product showed that they were conserved among the clinical A. actinomycetemcomitans isolates representing all currently known serotypes. AaPAL expression was shown under different nutritional and atmospheric conditions that resembled those in periodontal pockets. PAL deficiency in turn led to pleiotropic effects on the phenotype of A. actinomycetemcomitans, such as cell elongation and decreased growth rate. To purify AaPAL we employed affinity chromatography using anti-AaPAL peptide antibodies. The extensive characterization of the purified AaPAL by SDS-PAGE gel staining and mass spectrometry demonstrated that the final purification product did not contain other bacterial proteins than AaPAL. The protein had not lost its antigenicity during purification, since it was recognized by sera from patients with A. actinomycetemcomitans-associated oral and nonoral infections. AaPAL also appeared to be a strongly immunoreactive antigen in patients with periodontitis whose serum IgG antibodies recognized in western blot a 17 kDa OMP in the parental strain but not in the pal-deficient mutant. In addition to its immunogenicity, AaPAL also induced proinflammatory cytokine and chemokine response from human whole blood as determined by a cytokine antibody array. A cell culture insert model was designed to study how bacterial components could be introduced to the host in infections. The experiments demonstrated that live bacteria released extracellularly free-soluble AaPAL, but also other components, via an unknown outer membrane vesicle-independent mechanism. The immunogenicity and proinflammatory potential of the previously uncharacterized outer membrane lipoprotein of A. actinomycetemcomitans, AaPAL, suggests that it contributes to the pathogenicity of this bacterium. That live A. actinomycetemcomitans cells released free-soluble cell components may represent a new pathogenic mechanism.

<p>The aim of this study is to examine the knowledge and experience of the bad breath among high school students in Kristianstad. A further objective was to measure the amount of VSC (Volatile Sulfur Compounds) among those who perceive themselves to have a bad breath. A questionnaire with 21 closed questions was distributed to 120 high school students from Kristianstad municipality who were between the ages of 17-20. The results of this study revealed that most of the students had good knowledge of halitosis. The majority of all participants thought it was important to smell fresh in the mouth and also experienced most of their breath as very good or good. Only a few felt that they had bad breath. However, these persons did think their breath affect their everyday life. Furthermore, they have never felt embarrassed or caught in embarrassing situations due to their breath. The results from the clinical examination was carried out on 8 of the 120 students and was designed to measure the amount of sulphurous gases in the oral cavity which revealed that the average values that emerged in the examination below is the limited values for having a poor breath.</p> / <p>Syftet med denna studie var att undersöka kunskapen och upplevelsen av dålig andedräkt hos gymnasieelever i Kristianstad, ytterligare ett syfte var att mäta mängden VSC (reaktiva svavelföreningar)  hos dem som upplever sig ha dålig andedräkt. En enkät med 21 slutna frågor delades ut till 120 gymnasieelever som var mellan 17-20 år och studerade i Kristanstads kommun. Av resultatet i denna studie framkom att de flesta av gymnasieeleverna hade goda kunskaper om halitosis. Nästan alla 120 som besvarade enkäten tyckte att det var viktigt att lukta fräsch i munnen och dessutom upplevde de flesta sin andedräkt som mycket bra eller bra. Endast ett fåtal (14% ) kände att de ibland hade dålig andedräkt. Hos dessa  påverkades inte andedräkten deras vardagliga liv och de hade aldrig blivit generade eller hamnat i pinsamma situationer på grund av sin andedräkt. Resultatet från den kliniska undersökningen som gjordes på 8 av de 120 elever som besvarade enkäten där mängden av de svavelhaltiga gaserna i munhålan mättes visade att i genomsnitt ligger värden som framkom i undersökningen under gränsvärden för att man skall ha dålig andedräkt.</p>

The aim of this study is to examine the knowledge and experience of the bad breath among high school students in Kristianstad. A further objective was to measure the amount of VSC (Volatile Sulfur Compounds) among those who perceive themselves to have a bad breath. A questionnaire with 21 closed questions was distributed to 120 high school students from Kristianstad municipality who were between the ages of 17-20. The results of this study revealed that most of the students had good knowledge of halitosis. The majority of all participants thought it was important to smell fresh in the mouth and also experienced most of their breath as very good or good. Only a few felt that they had bad breath. However, these persons did think their breath affect their everyday life. Furthermore, they have never felt embarrassed or caught in embarrassing situations due to their breath. The results from the clinical examination was carried out on 8 of the 120 students and was designed to measure the amount of sulphurous gases in the oral cavity which revealed that the average values that emerged in the examination below is the limited values for having a poor breath. / Syftet med denna studie var att undersöka kunskapen och upplevelsen av dålig andedräkt hos gymnasieelever i Kristianstad, ytterligare ett syfte var att mäta mängden VSC (reaktiva svavelföreningar)  hos dem som upplever sig ha dålig andedräkt. En enkät med 21 slutna frågor delades ut till 120 gymnasieelever som var mellan 17-20 år och studerade i Kristanstads kommun. Av resultatet i denna studie framkom att de flesta av gymnasieeleverna hade goda kunskaper om halitosis. Nästan alla 120 som besvarade enkäten tyckte att det var viktigt att lukta fräsch i munnen och dessutom upplevde de flesta sin andedräkt som mycket bra eller bra. Endast ett fåtal (14% ) kände att de ibland hade dålig andedräkt. Hos dessa  påverkades inte andedräkten deras vardagliga liv och de hade aldrig blivit generade eller hamnat i pinsamma situationer på grund av sin andedräkt. Resultatet från den kliniska undersökningen som gjordes på 8 av de 120 elever som besvarade enkäten där mängden av de svavelhaltiga gaserna i munhålan mättes visade att i genomsnitt ligger värden som framkom i undersökningen under gränsvärden för att man skall ha dålig andedräkt.

Traditionally, bacterial culture has been used for bacterial detection, allowing study of living microorganisms. Molecular methods are rapid and allow simultaneous identification of numerous species and uncultivated phylotypes. The objective of this doctoral thesis was to investigate the role of the oral microbiota, including poorly characterized and uncultivated bacteria, in dental caries and periodontitis, by comprehensive molecular, clinical, and statistical methods. The microbiota of 275 pre-school children (75 with caries and 200 caries-free) was examined by whole genomic DNA probes, 16S rDNA cloning and sequencing, and PCR. Streptococcus mutans, exhibiting a combined association with Streptococcus sobrinus, was significantly associated with Early Childhood Caries (ECC). Plaque from children with Severe Early Childhood Caries (S-ECC) was diverse with 138 identified and 107 unidentified taxa, which possibly included novel phylotypes. Other species/phylotypes associated with childhood caries included Lactobacillus gasseri (p&lt;0.01), Lactobacillus fermentum, Actinomyces israelii, and Actinomyces odontolyticus (all p&lt;0.05, ECC), Veillonella parvula (p&lt;0.01), Veillonella atypica (p&lt;0.05), and Veillonella sp. HOT-780 (p&lt;0.01, S-ECC). Lactobacillus acidophilus and Lactobacillus reuteri, both used as probiotic therapy species, were detected more frequently in caries-free children than those with ECC. Fastidious periodontal species, including Parvimonas micra, Aggregatibacter actinomycetemcomitans, Eubacterium brachy, Filifactor alocis (all p &lt;0.05), and Porphyromonas gingivalis (p&lt;0.01), were also more frequently detected in children with dental caries than in caries-free children. Other variables associated with ECC were race, dental visit, snacking (all p&lt;0.05), and visible dental plaque (p&lt;0.01). The oral microbiota of early periodontitis in young adults (N=141) was analyzed by whole genomic and oligonucleotide DNA probes, and PCR. Species detected more frequently in early periodontitis than periodontal health included Treponema denticola, F. alocis, Porphyromonas endodontalis, Bacteroidetes sp. HOT-274 (oral clone AU126), and A. odontolyticus (p&lt;0.01) by oligonucleotide DNA probes, and P. gingivalis (p&lt;0.001) and T. forsythia (p=0.03) by PCR. Subgingival samples exhibited a higher prevalence of periodontitis-associated species than samples from tongue surface, including A. actinomycetemcomitans, T. denticola, T. forsythia (all p&lt;0.05), and uncultivated TM7, Treponema, and Actinobaculum clones (all p&lt;0.05). P. gingivalis (p&lt;0.01) by PCR was associated with periodontal disease progression. Early periodontitis was associated with older age (p=0.01), male gender (p=0.04), and cigarette smoking (p=0.05). The role of bacterial subgroups in periodontitis was examined by studying the serotypeability of 313 genotyped clinical A. actinomycetemcomitans isolates (189 subjects). A total of 95 strains (30 subjects) remained non-serotypeable, although PCR revealed presence of the serotype- specific genes. The absence of the immunodominant serotype-specific antigen was confirmed by immunoblot assays. No major DNA rearrangement in the studied serotype-specific gene clusters was found. In summary, detection of previously cultured species and uncultivated phylotypes revealed the diversity of the oral microbiota in dental diseases and health already early in life. Bacterial species have insufficiently characterized subgroups that may have attributes to evade the host response. Molecular approaches used in this study enable comprehensive, culture-independent characterization of the oral microbiome that may in the future lead to identification of diagnostic bacterial profiles for dental diseases.